This invention is in the field of plant molecular biology. More specifically, this invention pertains to nucleic acid fragments encoding cyclopropane synthetase in plants and seeds.
Plant oils and lipids are central to the plant membrane function and climatic adaptation and have a variety of nutritional and industrial uses. Production of oils with a modified fatty acid spectrum and improved functional characteristics is a major goal of the agricultural and food industry. Lipids represent a vast array of chemical structures many of which result, directly or indirectly, from metabolic processes that alter the degree of saturation (or unsaturation) and/or add functional groups to the lipids. These groups may be hydroxyls, ketones, epoxys, cyclopentenyl, cyclopropyl, furans or halogens. Using genetic engineering to change the activity of enzymes involved in the biosynthesis of lipids represents an attractive target for altering the levels of specific lipid structures in plants. Changes in the lipid profile will result in plants with superior oil qualities such as functional stability and/or taste. These plants will have considerable importance and value.
Cyclopropane-fatty-acyl-phospholipid synthase (EC 2.1.1.79) is also called cyclopropane synthetase or unsaturated-phospholipid methyltransferase. This enzyme adds a methylene group across the 9,10 position of a delta9-olefinic acyl chain in phosphatidylethanolamine or, more slowly, phosphatidylglycerol or phosphatidylinositol forming a cyclopropane deirvative. It transfers a methylene group from S-adenosyl-1-methionine to the cis double bond of the unsaturated fatty acid chain resulting in the replacement of the double bond with a methylene bridge. Cyclopropane synthetase from bacteria such as Escherichia coli and Mycobacterium tubeculosis has been isolated and their gene identified (Taylor, F. R. et al. (1981) Methods Enzymol 71:133-139; Grogan, D. W. and Cronan, J. E. Jr. (1984) J Bacteriol 158:286-295; Yuan, Y. et al. (1985) Proc Natl Acad Sci USA 92:6630-6634). No cycloprane synthetase has been identified to date in plants. Identification of the genes involved in the modification and saturation of lipids in plants is important for the development of industrially-important oil crops.
The instant invention relates to isolated nucleic acid fragments encoding cyclopropane synthetase. Specifically, this invention concerns an isolated nucleic acid fragment encoding a cyclopropane synthetase. In addition, this invention relates to a nucleic acid fragment that is complementary to the nucleic acid fragment encoding cyclopropane synthetase.
An additional embodiment of the instant invention pertains to a polypeptide encoding all or a substantial portion of a cyclopropane synthetase.
In another embodiment, the instant invention relates to a chimeric gene encoding a cyclopropane synthetase, or to a chimeric gene that comprises a nucleic acid fragment that is complementary to a nucleic acid fragment encoding a cyclopropane synthetase, operably linked to suitable regulatory sequences, wherein expression of the chimeric gene results in production of levels of the encoded protein in a transformed host cell that is altered (i.e., increased or decreased) from the level produced in an untransformed host cell.
In a further embodiment, the instant invention concerns a transformed host cell comprising in its genome a chimeric gene encoding a cyclopropane synthetase, operably linked to suitable regulatory sequences. Expression of the chimeric gene results in production of altered levels of the encoded protein in the transformed host cell. The transformed host cell can be of eukaryotic or prokaryotic origin, and include cells derived from higher plants and microorganisms. The invention also includes transformed plants that arise from transformed host cells of higher plants, and seeds derived from such transformed plants.
An additional embodiment of the instant invention concerns a method of altering the level of expression of a cyclopropane synthetase in a transformed host cell comprising: a) transforming a host cell with a chimeric gene comprising a nucleic acid fragment encoding a cyclopropane synthetase; and b) growing the transformed host cell under conditions that are suitable for expression of the chimeric gene wherein expression of the chimeric gene results in production of altered levels of cyclopropane synthetase in the transformed host cell.
An addition embodiment of the instant invention concerns a method for obtaining a nucleic acid fragment encoding all or a substantial portion of an amino acid sequence encoding a cyclopropane synthetase.